So far no Francisella tularensis proteins that can be used for induction of T cell protective immunity have been identified. The main goal of the proposed studies is to identify Francisella tularensis (Ft)-derived proteins that can serve for developing a safe, acellular vaccine against Ft. The Expression-Library Immunization (ELI) technique will be used to take advantage of the relatively small genome of Ft and the already established protocols for plasmid DNA immunization in rodents. Two separate representative genomic libraries will be created and tested. The first will encode secreted products that will be processed via the class II MHC presentation pathway and will stimulate CD4, CD8 T cell and humoral immunity. Alternatively, products from the second one will be targeted to the proteasome and processed via the class I MHC presentation pathway for primarily CD8 T cell stimulation. Screening of the libraries will be performed using an in vitro system that closely mimics in vivo Ft infection. Briefly, spleen cells from immunized or non- immunized animals are co-cultured for 6-8 hours with Schu S4 infected syngeneic bone marrow macrophages. Spleen cells are then stained for intracellular cytokines (glFN or IL-4) and surface determinants and analyzed by flow cytometry. Clones leading to development of anti-Ft abT cell immunity in mice will be collected and characterized.